Currently available techniques for detecting alternative pre-mRNA splicing rely on the use of fluorescent or enzymatic reporter systems which produce a single reporter protein (see, e.g., Wagner et al., Methods Mol. Biol., 257:29-46 (2004)). The single reporter construct is usually co-transfected with a reporter for a second protein as an expression control. Typically, an alternative cassette exon disrupts the translation of the reporter protein when included in the mature mRNA. The skipping of the alternative exon generates the functional mRNA and protein, while all other events (e.g., aberrant splicing, general inhibition of splicing, inhibition of transcription and translation) will abolish the expression of the reporter.
A major drawback of these reporter systems, however, is their inability to distinguish between general inhibition of splicing and selection of the particular alternative splicing events. In addition, the dynamic range of these reporter systems is relatively narrow. Such limitations make currently available reporter systems practical only for the detection of major changes in pre-mRNA splicing.
As such, there is a need in the art for reporter systems which are capable of detecting alternative pre-mRNA splicing with increased specificity, sensitivity, and versatility. The present invention satisfies this and other needs.